Characterization of the Hemolytic Activity of Mastoparan Family Peptides from Wasp Venoms.

Biologically active peptides have attracted increasing attention in research on the development of new drugs. Mastoparans, a group of wasp venom linear cationic α-helical peptides, have a variety of biological effects, including mast cell degranulation, activation of protein G, and antimicrobial and anticancer activities. However, the potential hemolytic activity of cationic α-helical peptides greatly limits the clinical applications of mastoparans. Here, we systematically and comprehensively studied the hemolytic activity of mastoparans based on our wasp venom mastoparan family peptide library. The results showed that among 55 mastoparans, 18 had strong hemolytic activity (EC50 ≤ 100 µM), 14 had modest hemolytic activity (100 µM < EC50 ≤ 400 µM) and 23 had little hemolytic activity (EC50 > 400 µM), suggesting functional variation in the molecular diversity of mastoparan family peptides from wasp venom. Based on these data, structure-function relationships were further explored, and, hydrophobicity, but not net charge and amphiphilicity, was found to play a critical role in the hemolytic activity of mastoparans. Combining the reported antimicrobial activity with the present hemolytic activity data, we found that four mastoparan peptides, Parapolybia-MP, Mastoparan-like peptide 12b, Dominulin A and Dominulin B, have promise for applications because of their high antimicrobial activity (MIC ≤ 10 µM) and low hemolytic activity (EC50 ≥ 400 µM). Our research not only identified new leads for the antimicrobial application of mastoparans but also provided a large chemical space to support the molecular design and optimization of mastoparan family peptides with low hemolytic activity regardless of net charge or amphiphilicity.

Promotion of HepG2 cell apoptosis by Sedum emarginatum Migo and the mechanism of action.

BACKGROUND: Sedum emarginatum Migo(S. emarginatum) has anti-tumor and anti-oxidant effects. This study aimed to screen the extractions of S. emarginatum against liver cancer in vitro and explore its anti-liver cancer mechanism. METHODS: The CCK-8(Cell Counting Kit-8) method was used to detect the inhibitory effect of different extracts of S. emarginatum on the proliferation of liver cancer HepG2 cells. The morphological changes of the cells after administration were observed with microscopy, cell apoptosis was detected by flow cytometry, and the expression of Bax, Bcl-2 and Caspase-3 mRNA in the cells were detected by RT-PCR (Reverse Transcription-Polymerase Chain Reaction) to explore the mechanism of action. RESULTS: CCK-8 method test results showed that among the different extracts of S. emarginatum, the ethyl acetate extract(1000 µg/ml, 2000 µg/ml, 2500 µg/ml, 3000 µg/ml) and n-butanol extract(1000 µg/ml, 2000 µg/ml, 2500 µg/ml, 3000 µg/ml) have the strongest inhibitory effect on the proliferation of HepG2 cells. In these 4 concentrations, the inhibitory effect increased as the concentration increased. The IC50 of the ethyl acetate extract on HepG2 cells was less than that of the n-butanol extract, so the ethyl acetate extract has a better proliferation inhibitory effect on HepG2 cells than the n-butanol extract, followed by the 70% ethanol extract(3000 µg/ml) and the water extract(3000 µg/ml), petroleum ether extract was the weakest. The results of microscopy showed that ethyl acetate extract caused hepatocarcinoma HepG2 cell morphology changed, cell density decreased, and suspension cells increased. Moreover, the results of flow cytometry showed that the ethyl acetate extract of S. emarginatum could induce HepG2 cell apoptosis at the concentrations of 2500µg/ml and 3000µg/ml. RT-PCR results showed that the expression of Bax mRNA was up-regulate by the middle(2500 µg/ml) and high(3000 µg/ml) dose groups of ethyl acetate extract. The expression of Caspase-3 mRNA was up-regulated by the low(2000 µg/ml), medium(2500 µg/ml) and high(3000 µg/ml) dose groups of ethyl acetate extract. The expression of Bcl-2 mRNA was down-regulated by the high(3000 µg/ml) dose group of ethyl acetate extract. CONCLUSION: The ethyl acetate extract of S. emarginatum has the best effect on human liver cancer HepG2 cells. Its anti-hepatocellular mechanism may be related to affect the expression of apoptosis genes (Bax, Bcl-2 and Caspase-3mRNA) and promote the apoptosis of liver cancer cells. It provided a reference for the research and development of drugs for the treatment of liver cancer.

Fingerprint and multi-index content determination of ethyl acetate extract of Sedum emarginatum Migo.

Sedum emarginatum Migo (Aoyejingtian) is a perennial succulent herb of the sedum genus in the family Crassulaceae, which has the fountion of treating furuncle, swelling and haematemesis, hematochezia, menorrhagia and hepatitis. Preliminary studies of our research group had showed that the ethyl acetate extract of Sedum emarginatum Migo could inhibit the proliferation of liver cancer HepG2 cells. The establishment of a reasonable and feasible quality evaluation method for the effective parts of Sedum emarginatum Migo can provide a scientific basis for the further development and utilization of Sedum emarginatum Migo. In this study, a multi-wavelength conversion method was used to establish high-performance liquid chromatography (HPLC) fingerprints of the ethyl acetate extract of Sedum emarginatum Migo, and the method was also used to simultaneously determine the gallic acid, protocatechuic acid, caffeic acid, and ferulic acid, isoquercitrin and luteolin in the ethyl acetate extract of Sedum emarginatum Migo. The similarity of the fingerprints of the ethyl acetate extract of Sedum emarginatum Migo from different origins and the content of 6 components were compared. The established method was simple, accurate, table and reliable, which could provide a fast, accurate and reliable method for comprehensive evaluation of the quality of Sedum emarginatum Migo.

Sanpao herbs inhibit development of atopic dermatitis in Balb/c mice.

OBJECTIVE: To explore the effects of SANPAOCAO (SPC), a compound traditional Chinese folk medicine, on chronic dermatitis/eczema in mice induced by 2, 4-dinitrochlorobenzene (DNCB). METHODS: Thirty-three Balb/c mice were randomly divided into a negative control group, a positive control group, a prednisolone treatment group, an SPC ethanol extract treatment group, a Cardiospermum halicacabum ethanol extract treatment group, a Physalis minima ethanol extract treatment group, and a Jussiaea repens ethanol extract treatment group. Mice in the six treatment groups had twenty-five microliters of 0.1% DNCB in acetone/olive oil (3: 1) applied to each side of their right ears and dorsal skin three times a week, over a 5 week period. They were treated with prednisolone or the various kinds of ethanol extract after each challenge. The weight difference between the two ears, pathological changes in the right ears, dermal inflammatory cell numbers, and total serum Ig E levels were used to assess the effects of the drugs. RESULTS: after the 5 weeks of challenges, the weight differences of the ears in the SPC group and the prednisolone group were significantly less than those in the other groups. There was evidence of significant suppression of the development of dermatitis, as determined by a histological examination and the serum Ig E levels. CONCLUSION: SPC has beneficial effects when used in the treatment of chronic dermatitis-eczema in mice.